Macrophages and dendritic cells (myeloid cells; MCs) function as immunologic sentinels, patrolling tissues to recognize and eliminate foreign invaders. Unfortunately, these cells can also respond to chronic diet-induced obesity (DIO) with inflammatory activation, accumulating in insulin-responsive tissues in a sterile inflammatory cascade implicated in the development of diabetes type 2. Emerging data support the concept that this response is instigated by lipids released from, for example, fat or liver cells, that are taken up by resident MCs, modulating their function to incite an inflammatory sequence in the tissue. My NIDDK-sponsored Clinician-Scientist Career Development Award (K08) explores the important biomedical question of how MCs contend with pro-inflammatory lipids, in particular saturated fatty acids (FAs). I have hypothesized that MCs can reduce toxicity from a rising tide of intracellular FAs by storing them as triacylglycerol (TG), preventing their flux into damaging cellular compartments and pathways. I have used genetics to manipulate TG storage capacity in murine MCs in order to modulate FA-induced inflammation in tissues and ameliorate metabolic diseases. I showed that MCs with increased expression of acyl CoA: diacylglycerol acyltransferase 1 (DGAT1), an enzyme involved in TG synthesis (aP2-Dgat1), have increased TG storage capacity and resistance to FA-induced inflammation (4). Remarkably, mice transplanted with aP2-Dgat1 bone marrow have less diet-induced inflammation and MC infiltration in fat tissue, and are protected from diet-induced insulin resistance (4). This proposl seeks to extend the impact of my current K08 on delineating the role of MCs in the progression of DIO to diabetes, and does so in two ways: A. Until now, our work has focused on DGAT1. However, DGAT1 is one of two known mammalian DGAT enzymes (DGAT2 is the other), and their combined action accounts for essentially all the TG formed in most cell types. Therefore, determining the degree to which TG storage per se limits the toxic effects of saturated FAs in MCs requires tools that target both DGAT enzymes, allowing the study of MCs in which TG storage capacity has been abolished altogether. We propose to transplant genetically modified hematopoietic progenitors that cannot store TG into irradiated mice, producing mice with MCs lacking TG storage capacity altogether. We will test the response of these mice to a chronically high-fat diet. B. Until now, we have focused on processes governing inflammation and insulin resistance in white adipose tissue. However, MCs coexist with parenchymal cells in several other tissues important to the development of diabetes, and we would like to explore how MC TG storage modulates inflammation in each of these contexts. We propose here to do so in the liver, by determining the role of MC-specific TG storage in the progression of fatty liver disease to liver inflammation and insulin resistance.